Host Responses to Coccidioides by Human Airway Epithelium

人体气道上皮对球孢子菌的宿主反应

• 批准号: 10373208
• 负责人: Jatin M Vyas
• 金额: $ 25.2万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-02 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10373208
• 关键词: Address; Animals; Antifungal Antibiotics; Apical; Architecture; Attenuated; CRISPR screen; Cell Communication; Cell Culture Techniques; Cell Line; Cells; Clustered Regularly Interspaced Short Palindromic Repeats; Coccidioides; Coccidioides immitis; Coccidioides posadasii; Complication; Critical Pathways; Data; Direct Costs; Disease; Dissection; Distal; Environment; Epithelial; Epithelial Cells; Extravasation; Foundations; Future; Gene Expression Profile; Genetic Transcription; Host Defense; Human; Human body; IL8 gene; Immune; Immune response; Immunity; Immunocompetent; Immunocompromised Host; Immunologics; Individual; Infection; Inflammation Mediators; Inflammatory; Inhalation; Innate Immune Response; Investigation; Knowledge; Lead; Lung; Lung infections; Mediating; Methodology; Methods; Modeling; Monitor; Mus; Mycoses; Natural Resistance; Pathogenicity; Pathway interactions; Patients; Phagocytosis; Play; Proteins; Response Elements; Sampling; Signal Pathway; Signal Transduction; Site; Soil; Sputum; Surface; System; T cell response; Tissues; Transgenic Organisms; United States; Work; airway epithelium; base; biosafety level 3 facility; chemokine; cytokine; dimorphism; experimental study; fungus; genome-wide; in vivo; innovation; insight; knockout gene; lentiviral-mediated; mortality; mouse model; new technology; novel; pathogen; pathogenic fungus; recruit; response; single-cell RNA sequencing; stem cells; transcriptome; transcriptomics; vaccine candidate

PROJECT SUMMARY Invasive fungal infections represent a major threat to immunocompromised patients and despite the availability of anti- fungal antibiotics, mortality rates remain as high as 50%. In the human host, the airway epithelium is the first point of contact upon inhalation of fungal conidia. As an immunologically active tissue, the airway epithelium may participate in active phagocytosis and coordinated immune cell recruitment. Soil fungi, including Coccidioides, do not need a human host to propagate, but upon disruption can enter the human body and lead to infection if not cleared. The fungal dimorph Coccidioides is native to arid regions in the southwestern U.S. and can establish infections in both immunocompetent and immunocompromised individuals. Unfortunately, little is known with regards to the initial response of human airway epithelium to Coccidioides. Currently, the Δcps1 strain of Coccidioides is being investigated as a potential vaccine candidate. Leveraging new technologies that permit the isolation of human airway stem cells, we demonstrated that primary differentiated human airway epithelial cells recapitulate the complexity of human airways. Our preliminary data showed infection at the apical surface of human airway epithelial cells by C. posadasii (Δcps1) elicits pro-inflammatory signals by the epithelium. Lastly, we demonstrated effective gene knockout in primary human airway epithelial cells using non-viral bulk nucleofection-based CRISPR strategy. Thus, our primary objective is to decipher the transcriptional and secretomal signatures critical to mount a successful response to Coccidioides in the human airway epithelium. To address our primary objective, we propose the following two specific aims: [1] determine the host response of human airway epithelium to both WT and the vaccine candidate C. posadasii (Δcps1), and [2] identify key signaling pathways to elicit an immune response to WT and the vaccine candidate C. posadasii (Δcps1) by human airway epithelium. This work will identify essential components of the human airway epithelium to mediate a protective response to these fungal infections and will provide the necessary foundation for future experiments to dissect the key pathways responsible for an effective host response to C. posadasii.

项目摘要 侵袭性真菌感染是免疫功能低下患者的主要威胁，尽管有抗真菌药物 真菌抗生素，死亡率仍高达50%。在人类宿主中，气道上皮细胞是第一点， 吸入真菌分生孢子后接触。作为一种免疫活性组织，气道上皮可能参与 主动吞噬作用和协调的免疫细胞募集。土壤真菌，包括球孢子菌，不需要人类宿主 传播，但一旦破坏，如果不清除，可以进入人体并导致感染。真菌的二型体 球孢子菌属原产于美国西南部的干旱地区，可以在免疫活性和 免疫力低下的人。不幸的是，关于人体气道的初始响应知之甚少 上皮到球孢子菌属。目前，球孢子菌属的Δ cps1菌株正在作为潜在的疫苗进行研究 候选人利用允许分离人类气道干细胞的新技术，我们证明了主要的 分化的人气道上皮细胞概括了人气道的复杂性。我们的初步数据显示 C.感染人气道上皮细胞的顶面。posadasii（Δ cps1）通过以下途径激发促炎信号： 上皮细胞最后，我们证明了使用非病毒载体在原代人气道上皮细胞中的有效基因敲除。 基于批量核转染的CRISPR策略。因此，我们的主要目标是破译转录和分泌 这些特征对于在人气道上皮中对球孢子菌成功应答至关重要。为了解决我们的首要问题 目的：[1]确定人气道上皮细胞对两种药物的反应， WT和候选疫苗C. posadasii（Δ cps1），并[2]确定引发免疫应答的关键信号通路 到WT和候选疫苗C。posadasii（Δ cps1）。这项工作将确定必要的 人呼吸道上皮细胞的组成部分，以介导对这些真菌感染的保护性反应，并将提供 必要的基础，为未来的实验，解剖的关键途径负责有效的主机响应C。 posadasii。